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A series of organic solutions with different cellulose concentrations are prepared by dissolving natural cotton fibers in lithium chloride/dimethyl acetamide (LiCl/DMAC) solvent system after the activation of cotton fibers. Under different coagulating bath, the regenerated cellulose membranes are formed in two kinds of coagulation baths, and two coating methods including high-speed spin technique (KW-4A spin coating machine) and low-speed scraping (AFA-Ⅱ Film Applicator) are selected in this paper. The macromolecular structure, mechanical properties, crystallinity, thermal stability and wetting property of the regenerated cellulose membrane are characterized by Scanning Electron Microscope(SEM), Fourier Transform Infrared Spectroscopy (FT-IR),X-ray diffraction (XRD), Thermogravimetric analysis (TG) and contacting angle tester. The effects of mass fraction, coagulation bath type, membrane forming process on the regenerated membrane properties are investigated. Experimental results show that the performance of regenerated cellulose membrane is relatively excellent under the condition of using the KW-4A high-speed spin method, water coagulation bath, and when mass fraction of cellulose is 3.5%. The crystallinity of the regenerated cellulose membrane changes a lot compared with natural cotton fibers. The variation trend of thermal stability is similar with that of cotton fiber. But thermal stability is reduced to some degree, while the wetting ability is improved obviously.